Enhanced conductometric immunoassay for hepatitis B surface antigen using double-codified nanogold particles as labels

Abstract

A new conductometric immunoassay for hepatitis B surface antigen (HBsAg) was developed based bioelectrocatalytic reaction on a microcomb-type electrode by using double-codified nanogold particles as labels. This microcomb-type electrode was fabricated on an interdigitated transducer covered with a well-ordered anti-HBs/protein A/nanogold architecture. The double-codified nanogold particles were prepared by using nanogold-labeled anti-HBs antibodies conjugated with horseradish peroxidase (HRP). Sandwich-type immunoassay protocol was successfully introduced for the detection of HBsAg. The formation of the immunocomplex changed the direct electrical communication between the carried HRP and the electrode, and thus local conductivity variations could be assayed based on the bioelectrocatalytic reaction of the carried HRP in 0.01M PBS (pH 7.0) containing 60μM H2O2, 0.08M KI and 0.1M NaCl. Under optimized conditions, the linear range obtained by using HRP-conjugated anti-HBs as secondary antibodies was 1.5–450ng/mL HBsAg, while the assay sensitivity by using double-codified nanogold particles could be further increased to 0.01ng/mL with the linear range from 0.1 to 600ng/mL HBsAg. The developed immunoassay method showed good precision, high sensitivity, acceptable stability and reproducibility, and could be used for the detection of real sample with consistent results in comparison with those obtained by the ELISA method.